Toxoplasma gondii is a ubiquitous parasite that can cause birth defects in pregnant women and neurological problems in immunocompromised patients. Although the definitive host of this parasite is the cat, T. gondii can infect virtually any nucleated cell. Recently, the apical membrane antigen-1 (AMA1) has emerged as a potentially important component of the parasite's invasion machinery. The predicted structure of AMA1, the time of its release onto the parasite surface and its location at the extreme apical end of the parasite suggest that it plays a role during host cell entry. The long-term objective of this proposal is to determine how AMA1 facilitates host cell infection by T. gondii. To meet the first specific aim, knock out and/or anti-sense technology will be used to determine whether AMA1 is an essential gene. Secondly, an AMA1 immunoadhesin will be constructed to establish whether the ectodomain of AMA1 has a binding function. This tool can then be used to identify the host cell ligand(s) for AMA1. Finally, a variation of the yeast two-hybrid system will be used to identify cytoplasmic partner(s) for the intracellular domain of AMA1. These studies should provide critical information on how Apicomplexan parasites attach to and invade mammalian cells.